Silicon phthalocyanine dyes with axial ligands were described by Kenney et al., in U.S. Pat. No. 3,094,536 as well as by Joyner, R. D. et al., J. Inorg. Nucl. Chem. 1960, 15, 387 and Esposito, J. N. et al., Inorg. Chem. 1966, 5, 1979–1984. The method of making silicon phthalocyanine is to react a phthalocyanine precursor e.g., diiminoisoindoline, with silicon tetrachloride to form silicon phthalocyanine directly. Recently, a more flexible method to make silicon phthalocyanine has been disclosed in U.S. Pat. No. 5,872,248, by inserting silicon into a metal-free phthalocyanine. The method includes providing a metal-free phthalocyanine and reacting the metal-free phthalocyanine with HSiCl3 to provide a reaction product, then reacting the reaction product with water; and extracting a silicon phthalocyanine.
A variety of silicon phthalocyanines have been reported. The application of silicon phthalocyanine dyes span wide fields, including for example, colorants, cancer therapy agents, detergents, and non-linear optical materials. However, silicon phthalocyanine dyes that are used as fluorescent reporter groups in bioassays are rare due to the fact that only a few water soluble or reactive silicon phthalocyanine dyes have been reported to date.
One silicon phthalocyanine dye having both water solubility and the activated/reactive functionalities, which is useful as a marker in bioassays, can be found in U.S. Pat. Nos. 6,060,598, 5,846,703, 5,403,928; and in WO 91/18007. These patent documents describe a silicon phthalocyanine dye structure, known as “La Jolla Blue” dye, which features two water-soluble axial polyoxyhydrocarbyl moieties, and also two reactive carboxylic acid groups on their peripheral positions. This dye is a mixture of similar dye structures due to the polymer groups attached.
Water solubility of dyes is a useful feature for bioassays. Even though only a few water-soluble silicon phthalocyanine dyes have been made, other phthalocyanine dye structures with water-solubility are known. These water-soluble phthalocyanine dyes typically have four identical water-soluble substituents on the peripheral positions of the macrocycle. Water soluble groups include phosphonate groups, carboxylate groups, sulfonate groups, quaternary ammonium groups or pyridinium groups (see, for example, Sharman, W. M. et al, Tetrahedron Lett. 1996, 37(33), 5831–5834; Wang, X. et al, Dyes and Pigments 1999, 41, 193–198; Ngai, T. et al, Langmuir 2001, 17, 1381–1383; Weber, J. H. et al, Inorg. Chem. 1965, 4(4), 469–471; Yang, Y. C. et al, Inorg. Chem. 1985, 24, 1765; Kimura, M. et al, J. of Porphyrins and Phthalocyanines 1997, 1, 309–313; Filippis, M. P. D. et al, Tetrahedron Lett. 2000, 41, 9143–9147; Chen, Z. et al, Langmuir 2001, 17, 7957–7959; Minnock, A. et al, Antimicrobial Agents and Chemotherapy 2000, 44(3), 522–527). These water soluble phthalocyanines are isomeric mixtures due to the random substitution of the water-soluble groups on the macrocycle, and do not have an activated or reactive group which can be utilized to conjugate biomolecules.
The presence of an activated/reactive group on the dye structure is another useful feature for a dye in bioassays. To make a mono-functional activated/reactive phthalocyanine dye, mono-substituted unsymmetrical phthalocyanine dye structures are desired for one class of dyes. However, mono- or di-substituted unsymmetrical phthalocyanines are difficult to synthesize. In most cases, mono- or di-substituted unsymmetrical phthalocyanines are transition metal or metal-free phthalocyanines without aqueous solubility (see, for example, Hu, M. et al., J. Med. Chem. 1998, 41, 1789–1802; Weitemeyer, A. et al., J. Org. Chem. 1995, 60, 4900–4904; Sastre, A. et al., J. Org. Chem. 1996, 61, 8591–8597; Sastre A. et al Tetrahedron Letters 1995, 36(46), 8501–8504; Kasuga, K. et al Inorganica Chimica Acta. 1992, 196, 127–128; and Kobayashi, N. et al., J. Am. Chem. Soc. 1990, 112, 9640–9641). Synthetic strategies for mono- or di-substituted phthalocyanines include a mixed condensation of two different dinitriles or diiminoisoindolines, or by a selective synthetic strategy to expand a subphthalocyanine with substituted diiminoisoindolines. The substituents on unsymmetrical phthalocyanines are typically non-reactive groups, but a few reactive mono- or di-substituted unsymmetrical phthalocyanines, such as monohydroxyl zinc or metal-free phthalocyanines (see, Hu, M. et al J. Med. Chem. 1998, 41, 1789–1802) and monoaminated zinc or metal-free phthalocyanines (see, Sastre A. et al Tetrahedron Letters, 1995, 36(46), 8501–8504) have been made. These reactive groups on the phthalocyanine dyes have low reactivity, are not water-soluble, and are difficult to derivatize.
A group of phthalocyanine dyes and their analogues were described in U.S. Pat. Nos. 5,346,670 and 5,135,717. However, these phthalocyanine dyes are isomeric mixtures due to the random substitution of the water-soluble groups.
In view of the foregoing shortcomings, what is needed in the art are new phthalocyanine dyes that are water soluble, isomericly pure, and which possess high quantum yield. The present invention satisfies these and other needs.